The studies reported here demonstrate some of the factors affecting the binding of benzo(a)pyrene (BP) to macromolecules in cultured human bronchial mucosa. Bronchial specimens were obtained at either surgery or "immediate" autopsy from patients with and without lung cancer. Grossly normal-appearing pieces of bronchus were cultured in a chemically defined medium. After 7 days, explant cultures were exposed to (H3) BP, usually for 24 hr, and then binding to total cellular macromolecules was studied by autoradiography, and binding to DNA was measured following isolation of DNA from bronchial mucosal cells. The extent of binding of (H3) BP was dependent on dose of BP, length of exposure to (H3) BP and temperature. By autoradiography, bronchial epithelial cells bound more (H3) BP than stromal fibroblasts. Both 7,8-benzoflavone and butylated hydroxytoluene appeared to reduce the level of (H3) BP bound to DNA, while nicotine apparently did not alter the level of binding. These studies demonstrate that the bronchial mucosa, an important human cancer target tissue, has the capability to form metabolites of BP which bind to macromolecules including DNA. In addition, 7,8-benzoflavone and butylated hydroxytoluene, both known to alter the microsomal metabolism of BP, reduce the level of (H3) BP bound to DNA.